Effects of magnetic nanoparticle of Fe3O4 on apoptosis induced by Gambogic acid in U937 leukemia cells / 中国实验血液学杂志

This study was aimed to explore the potential therapy of Gambogic acid (GA) combined with magnetic nanoparticle of Fe3O4 (Fe3O4-MNP) on leukemia. The proliferation of U937 cells and the cytotoxicity were evaluated by MTT assay. Cell apoptosis was observed and analyzed by microscopy and flow cytometry respectively. The expressions of gene and protein were detected by quantitative real-time polymerase chain reaction and Western blot respectively. The results showed that GA enhanced the cytotoxicity for U937 cells in dose- and time-dependent manners. The Fe3O4-MNP itself had not cytotoxicity, but could enhance the inhibitory effect of GA on proliferation of U937 cells. The apoptotic rate of U937 cells induced by combination of GA with Fe3O4-MNP was higher than that by GA alone. The typical apoptotic features of cells treated with GA and Fe3O4-MNP were observed. The expression levels of caspase-3 and bax after co-treatment of GA and Fe3O4-MNP were higher than that exposed to GA or Fe3O4-MNP alone, but the expressions of bcl-2, NF-kappaB and survivin were down-regulated. It is concluded that Fe3O4-MNP can promote GA-induced apoptosis in U937 cells, and the combination of GA with Fe3O4-MNP may be a safer and less toxic new therapy for leukemia.

MDR reversal activity of bromotetrandrine in vitro and in vivo / 中国实验血液学杂志

The present study was aimed to evaluate the MDR reversal activity of bromotetrandrine (BrTet) in vitro and in vivo. The inhibitory effects of adriamycin (ADM) used alone or in combination with BrTet or Tet on the proliferation of K562 and K562/A02 cells were evaluated by MTT assay. The ADM accumulation and the protein levels of P-glycoprotein (P-gp) were detected by flow cytometry. The mRNA levels of P-gp were determined by RT-PCR. The in vivo effect of BrTet and Tet was investigated by using nude mice grafted with sensitive human leukemia cell line K562 and MDR cell line K562/A02. The results showed that BrTet at 0.25, 0.5 and 1 micromol/L reversed the resistance to ADM in MDR K562/A02 cells in a dose-dependent manner. Flow cytometry suggested that BrTet significantly increased the intracellular accumulation of ADM in K562/A02 cells in a dose-dependent manner. BrTet also inhibited the overexpression of P-gp in K562/A02 cells, and down-regulated mdr1 expression. In nude mice bearing K562 xenografts on the left flank and K562/A02 xenografts on the right flank, intraperitoneal injection of 10 mg/kg BrTet significantly enhanced the antitumor activity of ADM against K562/A02 xenografts with inhibitory rates of 26.1%, while ADM alone inhibited the growth of K562/A02 xenografts only by 5.8%. No enhancement effect by BrTet was seen in K562 xenografts. It is concluded that BrTet shows significant MDR reversal activity in vitro and in vivo. Its activity may be related to the inhibition of P-gp overexpression and the increase intracellular accumulation of anticancer drugs. BrTet may be a promising-MDR modulator for eventual assessment in the clinic.

Effect of tetrandrine in combination with droloxifen on the expression of NF-kappaB protein in K562 and K562/A02 cell lines / 中华血液学杂志

<p><b>OBJECTIVE</b>To study the effect of tetrandrine (Tet) in combination with droloxifen (DRL) on the expression of nuclear factor kappa B (NF-kappaB) in K562 and K562/A02 cell lines and its reversal mechanism.</p><p><b>METHODS</b>The activation of NF-kappaB in K562 and K562/A02 cell lines and the effect of Tet or DRL alone or in combination on NF-kappaB protein expression were determined with immunocytochemistry and Western blotting respectively.</p><p><b>RESULTS</b>(1) K562/A02 cells displayed higher level of NF-kappaB protein expression than K562 cells. (2) The application of Tet or DRL alone or in combination had no effect on NF-kappaB expression in K562 cells at 6 h and 12 h (P > 0.05). (3) Tet and DRL alone or in combination could significantly down-regulate NF-kappaB protein expression in nuclei of K562/A02 cells. The effect was more significant in combination than either alone. This effect was more significant at 12 h than at 6 h.</p><p><b>CONCLUSIONS</b>(1) Activation of NF-kappaB may be involved in the mechanism of MDR of K562/A02 cell line. (2)Inhibition of NF-kappaB activation may be involved in the reversal of multidrug resistance in K562/A02 cells by Tet and DRL.</p>

Quantitative microarray-based DNA methylation analysis of E-cadherin gene promoter in acute leukemia / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To quantitatively detect the methylation of E-cadherin gene 5'-CpG islands in acute leukemia by microarray-based DNA analysis and to briefly discuss the role of microarry for detection of methylation in tumors.</p><p><b>METHODS</b>Bisulfite-modified DNA was used as a template for PCR amplification, resulting in conversion of unmethylated cytosine, but not methylated cytosine, into thymine within CpG islands of interest. Five sets of oligonucleotide probes were designed to fabricate a DNA microarray to detect the methylation changes of E-cadherin gene CpG islands in acute leukemia. By drawing a standard curve to assess the levels of changes in methylation detected in the examined samples.</p><p><b>RESULTS</b>Microarray assay was successfully used to quantitatively detect methylation changes of E-cadherin gene in 5 acute leukemia samples. Varying degree of methylation was detected in five regions and the hypermethylation region was the same. The result was validated by gene sequencing.</p><p><b>CONCLUSION</b>Microarray assay may be applied as an useful tool for mapping methylation changes in multiple CpG loci and for leukemia research. It is more time-saving and labor-saving than gene sequencing and can be used to quantitatively detect changes in methylation with high throughput.</p>

Detection of complex karyotype in a myelodysplastic syndrome cell line (MUTZ-1) by metaphase fluorescence in situ hybridization / 中国实验血液学杂志

This study was aimed to investigate the cytogenetic changes of MDS cell line (MUTZ-1) with chromosome 5q deletion. R-banding analysis was used to identify chromosome aberrations in MDS cell line and Vysis Spectra Vysion M-FISH was used to further characterize chromosomal complex karyotype. The results indicated that M-FISH exhibited obvious chromosomal aberrations with high frequency including translocation, insertion, breakage and rearrangement, deletion and increasement of chromosome number, the complex karyotype of MUTZ-1 was shown as 50, xx, der (1) t (1;2), ins (1;14), +der (2) t(2;19), der (3) t (3;5), der (3) (3::5::22), 5q-, der (6) t (3;6), der (7) (18::7::17), +8, +der (9) t (1;9), der (10) t (1;10), +11, +12, der (?13) (10::13::5::8), der (14) t (8;14), der (14) t (14, 15), der (15) t (15;21) x 2, +17, +18, -21, -22. It is concluded that M-FISH analysis revealed obvious changes in complex karyotype of MDS cell line MUTZ-1, and the M-FISH technique can increase accuracy of detection for chromosomal complex karyotype, and help diagnosis and prognostic evaluation of MDS.

Effects of sensitized donor lymphocyte infusion on the chimerism and graft-versus-host disease after nonmyeloablative allogeneic stem cell transplantation / 中国实验血液学杂志

To explore whether the complete donor chimerism could be achieved and graft-versus-host disease could be alleviated by donor lymphocyte infusion which was sensitized by the skin of the recipient, female C57BL/6 mice (H-2(b), B6) as recipients received total body irradiation (TBI) of 5.5 Gy ((60)Co gamma-ray) on day 0 followed by allogeneic hematopoietic stem cell transplantation (allo-HSCT). The allo-grafts consisted of 2 x 10(7) peripheral hematopoietic stem cells from mobilized male BALB/c (H-2(d)) donor mice with the granulocyte colony-stimulating factor (G-CSF). Day 2 after allo-HSCT, the recipient mice were given 200 mg/kg cyclophosphamide intraperitoneally. Afterwards these recipient mice were infused 2 x 10(6) sensitized or unsensitized-donor lymphocytes at the 28 days after transplantation. The results showed that the mice receiving sensitized-donor lymphocyte infusion did not suffer from GVHD and the phenotypic character of the recipient mice (black color) converted to that of the donor mice (white color), and to become full-donor chimerism. It was found that the ratio of CD4(+)/CD8(+) T lymphocytes of them decreased at the earlier period and increased after half month, but which were also lower than that of the normal value. While various grades of acute GVHD was observed in that of the control group and the mixed-chimeras were maintained, though it increased a little, and the ratio of CD4(+)/CD8(+) T lymphocytes increased at first, then decreased to the normal level half month later. It is concluded that sensitized DLI converted mixed to complete donor chimerism without GVHD, and the rate of CD4(+)/CD8(+) has close relation to the incidence of GVHD.